Glucose and amino acids evoke a complex series of cationic and metabolic events resulting in the activation of insulin secretions. The coupling between metabolic and cationic events appears to be due to the metabolic generation of protons (H+). H+ may serve a primary role in controlling the alterations of K+ and Ca++ permeabilities underlying the rhythmicity of the electrical events. In addition, changes in pH, may be the physiological regulator of cell-to-cell coupling in the islet as has been found to occur in other cells or tissues. Our specific hypothesis is twofold: 1) nutrient-induced changes in intracellular pH (pHi) play a role in triggering or modulating the oscillatory nature of the electrical activity in the B-cell plasma membrane and cell-to-cell coupling, and 2) that there must be efficient pH regulatory mechanisms in the plasma membrane for effective back regulation of pHi following alterations in glucose metabolism. We will examine the influence of changes in pH on electrical activity by changing medium pH and using permeable weak acids and bases or ionophores in the absence or presence of D-glucose, L-leucine, or Alpha-ketoisocaproic. To determine the extent to which H+ influences the oscillatory nature of the electrical events, we will alter pHi in the presence of agents known to inhibit or activate Ca++ and voltage-sensitive K+ conductances. The role of Na:H and HCO3:Cl exchange systems in regulating pHi will be examined by alteration of the ionic concentration gradient or by the addition of inhibitors of these transport systems. We will use NH4Cl and high PCO2 to alkalinize and acidify the cell interior, respectively, in the absence and presence of glucose to determine whether pHi influences cell-to-cell coupling of B-cells, as assessed by electrical and dye (Lucifer Yellow) coupling techniques in intact islets and in monolayer cultures. In order to measure the actual extent and time-related changes in pHi under the above experimental conditions parallel experiments will be conducted using pH-sensitive microelectrodes. These studies will provide evidence as to the extent to where H+ serve as a regulatory signal in intra- and intercellular events via the generation of voltage signals. Clarification of the mechanisms by which pHi influences the B-cell consequently insulin release may provide the basis for understanding the derangements in glucose metabolism associated with pH alterations in various physiological and pathological states.